This application relates to the field of antiviral compounds, specifically porphyrin and porphyrin-like compounds, such as phthalocyanines and chlorins, for the inhibition of infection and replication of human immunodeficiency virus.
AIDS, or acquired immunodeficiency disease, is characterized by an imbalance in two basic types of immune system cells, helper/inducer T lymphocytes and suppressor T lymphocytes, with the ratio of suppressor cells to helper/inducer cells greatly elevated. Helper/inducer T cells, defined by a surface antigen called CD4, are responsible for the induction of most of the functions of the human immune system, including the humoral immune response involving the production of antibodies by B lymphocytes and the cell-mediated response involving stimulation of cytotoxic T cells. A condition associated with HIV is AIDS-related complex, or ARC. Most patients suffering from ARC eventually develop AIDS.
Two related retroviruses can cause AIDS, human immunodeficiency virus type 1 and type 2 (HIV-1 and HIV-2, generally referred to herein as HIV). The genomes of the two viruses are about 50% homologous at the nucleotide level, contain the same complement of genes, and appear to attack and kill the same human cells by the same mechanism. Also known as LAV (lymphadenopathy-associated virus), HTLV-3 (human T-lymphotropic virus-type 3), and ARV (AIDS-related virus), HIV-1 was identified in 1983. Virtually all AIDS cases in the U.S. are associated with HIV-1 infection. HIV-2 was isolated in 1986 from West African AIDS patients.
Both types of HIV are retroviruses, in which the genetic material is RNA rather than DNA. The viruses carry with them a polymerase (reverse transcriptase) that catalyzes transcription of viral RNA into double-helical DNA. The viral DNA can exist as an unintegrated form in the infected cell or be integrated into the genome of the host cell. As presently understood, the HIV enters the T4 lymphocyte where it loses its outer envelope, releasing viral RNA and reverse transcriptase. The reverse transcriptase catalyzes synthesis of a complementary DNA strand from the viral RNA template. The DNA helix then inserts into the host genome where it is known as the provirus. The integrated DNA may persist as a latent infection characterized by little or no production of virus or helper/inducer cell death for an indefinite period of time. When the viral DNA is transcribed and translated by the infected lymphocyte, new viral RNA and proteins are produced to form new viruses that bud from the cell membrane and infect other cells.
No treatment capable of preventing or reversing the immunodeficiency of AIDS or ARC is currently available. All patients with opportunistic infections and approximately half of all patients with Kaposi's sarcoma have died within two years of diagnosis. Attempts at reviving the immune systems in patients with AIDS have been unsuccessful.
A number of compounds have apparent antiviral activity against this virus, including HPA-23, interferons, ribavirin, phosphonoformate, ansamycin, suramin, imuthiol, penicillamine, carbovir, 3'-azido-3'-deoxythymidine (AZT), and other 2',3'-dideoxy-nucleosides, such as 2',3'-dideoxycytidine (DDC), 2',3'-dideoxyadenosine (DDA), 2',3'-dideoxyinosine (DDI), 3'-azido-2',3'-dideoxyuridine (AzddU), 2',3'-dideoxy-2',3'-didehydrocytidine (D4C), 3'-deoxy-2',3'-didehydrothymidine (D4T) and 3'-azido-5-ethyl-2',3'-dideoxyuridine (AedU).
Inhibitors of cellular processes will often limit viral replication. Unfortunately, they are also usually toxic for the host and therefore cannot be prescribed for a prolonged period of time because of their toxicity. Although AZT is the drug of choice at this time for the treatment of AIDS, preliminary results indicate that AZT exhibits toxicity in a clinical setting, causing bone marrow suppression, resulting in anemia and neutropenia. See Yarchoan et al., Lancet 575-580 (1986).
Efforts to decrease the problem of toxicity have primarily been directed towards finding selective, less toxic drugs. Due to the exorbitant cost of the nucleoside type drugs, research has also been directed towards compounds which are relatively easy and economical to manufacture.
Photodynamic therapy (PDT) is the treatment of malignant tumors with photosensitizers, such as porphyrins and phthalocyanines. Briefly, certain photosensitizers, including porphyrins, metalloporphyrins, and phthalocyanines, localize preferentially in tumor cells. Irradiation of the tissue results in selective cell death of the cells carrying the photosensitizer. Red light in the therapeutically useful range of 600-1200 nm is used. Light in this region of the spectrum has increased transmittance in biological tissue. Both porphyrins and the structurally similar phthalocyanines absorb red light. While porphyrins have been studied more extensively, phthalocyanines have improved absorbance properties and higher extinction coefficients in this region of the spectrum.
General reviews of photodynamic therapy are by Gomer, C. J., et al., "Proceedings of the Clayton Foundation Conference on Photodynamic Therapy." Photochem. Photobiol. 46, 561-952 (1987) and Dougherty, T. J., Photochem. Photobiol. 45, 879-889 (1987).
The photochemistry and photophysics of porphyrins, metalloporphyrins, and phthalocyanines have been studied in detail. Processes observed include radiationless decay to ground, loss of an axial ligand, energy transfer, electron transfer, formation of singlet oxygen, phosphorescence and fluorescence. The photoprocesses observed in each system depend greatly on the central ligand, normally a metal (2H for porphyrin), the oxidation state of the metal and the axial ligand bound to the metal. A weaker dependence of the photophysical properties on the nature of the macrocycle is observed.
Porphyrins and phthalocyanines have been reported to have a variety of other biological activities. However, relatively little has been done with them with respect to in vivo clinical applications other than in photodynamic tumor therapy.
Perlin, et al., Antiviral Res. 7,43-51 (1987), recently reported that, upon exposure to light, hematoporphyrin, at concentrations as low as 0.5 .mu.g/ml, inhibits in vitro replication of influenza A and herpes simplex viruses, but not several other viruses. See also Lewin, et al., Proc. Soc. Exper. Biol. Med. 163, 81-90 (1980) and Schnipper, et al., J. Clin. Invest. 65, 432-438 (1980). Hematoporphyrin in combination with visible light also inhibits reverse transcription in vitro by the RNA-dependent DNA polymerase of Moloney leukemia virus from an exogenous template, as described by Munson, et al., Res. Commun. Chem. Pathol. Pharmacol. 16,175-178 (1977). Inhibition does not occur in the absence of exposure of the cells or viruses to light.
Within the last few months, hemin, ferric chloride protoporphyrin IX, has been shown to exhibit selective antiviral activity under certain conditions. Tsutsui and Mueller demonstrated in Biochem.Biophys.Res.Com. 149(2),628-634 (December 1987) that the reverse transcriptase activity of Rauscher murine leukemia virus, but not of avian myeloblastosis virus, was inhibited by hemin at a concentration of 10.sup.-4 M. They proposed that the hemin inhibited the reverse transcriptase activity by reversible, non-covalent interaction with the enzyme, not the RNA template.
Bhattacharya, et al., Proc. Natl. Acad. Sci. USA 78(5),2683-2687 (1981) showed that hemin differentially inhibits three forms of DNA polymerase .alpha., supporting the theory that hemin may be an important modulatory protein. None of these researchers correlated inhibition with toxicity, however, nor examined the compounds as selective antiviral drugs.
Ben-Hur, et al., Photochem. and Photobiol., 46(5):651-656, (1987) demonstrated that the incorporation of thymidine, uridine, and leucine, into DNA, RNA, and protein respectively, is inhibited in log-phase Chinese hamster cells that are photosensitized with chloroaluminium phthalocyanine tetrasulfonate. There are no known reports demonstrating that phthalocyanines possess antiviral activity.
It is therefore an object of the present invention to provide compounds having selective activity against Human Immunodeficiency Virus.
It is a further object of the present invention to provide compounds having antiviral activity which are relatively non-toxic.
It is a still further object of the present invention to provide compounds having antiviral activity which are relatively inexpensive and easy to manufacture.